Delineating the Rules for Structural Adaptation of Membrane-Associated Proteins to Evolutionary Changes in Membrane Lipidome.

Maria Makarova,Attila Glatz,Gabor Balogh,Paula Booth,Maria Peter,Snezhana Oliferenko,Nestor Lopez Mora,James I Macrae,Laszlo Vigh,Eugene Makeyev

doi:10.1016/j.cub.2019.11.043

Abstract

SummaryMembrane function is fundamental to life. Each species explores membrane lipid diversity within a genetically predefined range of possibilities. How membrane lipid composition in turn defines the functional space available for evolution of membrane-centered processes remains largely unknown. We address this fundamental question using related fission yeasts Schizosaccharomyces pombe and Schizosaccharomyces japonicus. We show that, unlike S. pombe that generates membranes where both glycerophospholipid acyl tails are predominantly 16–18 carbons long, S. japonicus synthesizes unusual “asymmetrical” glycerophospholipids where the tails differ in length by 6–8 carbons. This results in stiffer bilayers with distinct lipid packing properties. Retroengineered S. pombe synthesizing the S.-japonicus-type phospholipids exhibits unfolded protein response and downregulates secretion. Importantly, our protein sequence comparisons and domain swap experiments support the hypothesis that transmembrane helices co-evolve with membranes, suggesting that, on the evolutionary scale, changes in membrane lipid composition may necessitate extensive adaptation of the membrane-associated proteome.

Highlights

We detected some variation in the abundance of the minor GPL classes, sphingolipids, sterols, and storage lipids, with S. japonicus containing less sterols and sphingolipids (Figures 1C, 1D, and S1A; Table 1 in Data S1)
When supplemented together with C18:1, C10:0 was 4.6 times more efficiently incorporated into GPLs of cerulenin-treated S. japonicus as compared with S. pombe (Figure S1E; Table 5 in Data S1). These results demonstrate that S. japonicus physiology relies on the presence of both C10:0 and long-chain fatty acids (LCFAs)
Increasing temperature resulted in the growth of liquid disordered (Ld) domains in S. japonicus-derived membranes but phase coexistence was maintained at 37C, which is at the higher end of the physiologically relevant temperature range (Figure 2B). This was in spite of the lower sphingolipid to GPL ratio in this organism, as compared to S. pombe (Figure 1C). These results demonstrate that the divergence in GPL structures between the two fission yeasts may have a profound effect on the biophysical properties of their membranes, resulting in higher membrane stiffness and a distinct phase behavior in S. japonicus

Summary

Methods

Molecular genetics manipulations were performed using PCR- or plasmid-based homologous recombination using plasmids carrying S. japonicus ura, kanR, hygR or natR cloned into the pJK210-backbone (pSO550, pSO820 and pSO893, respectively). Using similar strategy and the construct carrying the hygR cassette as a selection marker, we replaced the fas ORF. We have identified several mis-annotations in the S. japonicus gene encoding the FAS b subunit (SJAG_04107). Using both RNaseq / de novo transcript assembly and sequencing of cDNA obtained from vegetatively grown cells we identified the following errors: 1) T at position 1240 is absent; 2) an additional C is present at position 2158; 3) both annotated introns are retained. A PCR-based method [79] was used to tag Anp, Gmh, Lro and Tcb at the C terminus, typically using KanMX6 or natR as a selection marker

Results

Discussion

Conclusion